Rapid functional annotation of native enhancers in embryonic stem cells with a Neisseria meningitidis CRISPR-Cas9 system
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Barbara Tabak
PROVIDER: E-GEOD-64059 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Shared Molecules
Only show the datasets with similarity scores above: 0.5
Threshold
0.5
Similar Datasets
Project description:Gene regulatory elements such as enhancers have profound effects on cellular function, health, and disease. Our understanding of mammalian enhancer function is limited by the lack of a technology that would allow for a rapid and thorough test of their cell type-specific function. Here, we describe a novel Cas9-effector system that enables rapid testing and functional annotation of native enhancers in embryonic stem cells. Total RNA obtained from R26 dCas9-effector mESC lines after viral delivery of sgRNAs directed against proximal promoter or enhancer regions
2015-03-03 | E-GEOD-64057 | biostudies-arrayexpress
Project description:Anterior foregut endoderm (AFE) gives rise to many tissue types of interest for therapeutic research including the esophagus, salivary glands, lung, thymus, parathyroid and thyroid. Despite its importance, only few reports describe the generation of AFE from pluripotent stem cells (PSCs) by directed differentiation. Here, we describe a novel protocol to derive a subdomain of AFE, identified by expression of Pax9, from PSCs using small molecules and chemically defined conditions. Generation of a reporter PSC line allows isolation and characterization of Pax9+ AFE cells. When transplanted in vivo, Pax9+ AFE can form several distinct types of complex anterior foregut epithelia including mucosal glands and stratified squamous epithelium. Finally, we show that the directed differentiation protocol can be used to generate AFE from DiGeorge Syndrome patient-specific human induced PSCs, thus creating a platform to produce anterior foregut derivatives for therapy and to enable the study of disorders of the AFE. Total RNA obtained from FACS purified from in vitro dervied mouse definitive endoderm, anterior foregut and ES cells. AFE cells were derived from a 129X1/SvJ background, DE cells from 129X1/SvJ x 129S1/SV-+p+Tyr- cKitlSl-J/+ (R1 ES cells) and non reporter ES cells from a 129P2/OlaHsd background.
2013-04-15 | E-GEOD-42139 | biostudies-arrayexpress
Project description:CKD and hypertension to impact a staggering 1.5 billion individuals within the next decade. Optimal fetal growth and development are outcomes of a delicate interplay between genetic and environmental factors. Nutrition emerges as a pivotal environmental determinant, orchestrating proper organogenesis. Malnutrition disrupts normal embryo development and potentially leads to chronic diseases in later life. Our understanding of the specific metabolic routes and their impact on kidney development is still elusive. Here, we used a multi-omics approach to study the importance of glucose metabolism to proper kidney development. We cultured E13.5 embryonic kidneys in the presence or absence of partial inhibition of glycolysis and submitted it to transcriptomic and proteomic profiling. We found that glycolysis-derived acetyl-CoA is an intracellular pleiotropic agent pivotal for proper kidney development.
2023-09-18 | PXD045445 | Pride
Project description:In this study we identify Mettl3, an m6A RNA modification writer, as a critical regulator for terminating naïve pluripotency and a positive maintainer of primed pluripotency in vitro and in vivo. Remarkably, Mettl3 knockout pre-implantation epiblasts and naïve ES cells, entirely lack m6A on coding mRNAs and are viable. Yet, they fail to adequately terminate the naïve pluripotent state, and subsequently undergo aberrant priming and early lineage commitment at the post-implantation stage. A comprehensive functional and genomic analysis involving profiling of m6A, RNA transcription and translation in Mettl3 wild-type and knockout pluripotent and differentiated cells, identified m6A as a critical determinant that destabilizes secondary naïve specific pluripotency genes Esrrb, Klf4 and Nanog, and restrains their transcript stability and translation efficiency. In summary, our findings provide for the first time evidence for a critical role for an mRNA epigenetic modification in early mammalian development in vivo, and identify a mechanism that functionally regulates mouse naïve and primed pluripotency in an opposing manner. polyA RNA-seq was measured in mouse embryonic stem cells (ESCs) and embroid bodies (EBs), each in WT and in Mettl3-KO cell lines. RNA-seq was measured also from WT mouse embronic fibroblasts (MEF). 3 biological replicates are available from ESCs and 2 from EBs. Replicate C in ESCs was measured alongside protein levels (SILAC) and was used for the analysis of that assay.
2014-11-02 | E-GEOD-61997 | biostudies-arrayexpress
Project description:miRNA sequencing in mouse embryonic stem cells according to miR-142 reporter activity
2015-12-31 | E-MTAB-2831 | biostudies-arrayexpress
Project description:In this study we identify Mettl3, an m6A RNA modification writer, as a critical regulator for terminating naM-CM-/ve pluripotency and a positive maintainer of primed pluripotency in vitro and in vivo. Remarkably, Mettl3 knockout pre-implantation epiblasts and naM-CM-/ve ES cells, entirely lack m6A on coding mRNAs and are viable. Yet, they fail to adequately terminate the naM-CM-/ve pluripotent state, and subsequently undergo aberrant priming and early lineage commitment at the post-implantation stage. A comprehensive functional and genomic analysis involving profiling of m6A, RNA transcription and translation in Mettl3 wild-type and knockout pluripotent and differentiated cells, identified m6A as a critical determinant that destabilizes secondary naM-CM-/ve specific pluripotency genes Esrrb, Klf4 and Nanog, and restrains their transcript stability and translation efficiency. In summary, our findings provide for the first time evidence for a critical role for an mRNA epigenetic modification in early mammalian development in vivo, and identify a mechanism that functionally regulates mouse naM-CM-/ve and primed pluripotency in an opposing manner. m6A-seq was measured from total RNA in mouse embryonic stem cells (ESCs), embroid bodies (EBs) and embronic fibroblasts (MEF). 3 biological replicates are available from BVSC ESC line and EBs, and two biological replicates are available for MEFs. Each sample consist of IP to m6A and control input
2014-11-02 | E-GEOD-61995 | biostudies-arrayexpress
Project description:In this study we identify Mettl3, an m6A RNA modification writer, as a critical regulator for terminating naïve pluripotency and a positive maintainer of primed pluripotency in vitro and in vivo. Remarkably, Mettl3 knockout pre-implantation epiblasts and naïve ES cells, entirely lack m6A on coding mRNAs and are viable. Yet, they fail to adequately terminate the naïve pluripotent state, and subsequently undergo aberrant priming and early lineage commitment at the post-implantation stage. A comprehensive functional and genomic analysis involving profiling of m6A, RNA transcription and translation in Mettl3 wild-type and knockout pluripotent and differentiated cells, identified m6A as a critical determinant that destabilizes secondary naïve specific pluripotency genes Esrrb, Klf4 and Nanog, and restrains their transcript stability and translation efficiency. In summary, our findings provide for the first time evidence for a critical role for an mRNA epigenetic modification in early mammalian development in vivo, and identify a mechanism that functionally regulates mouse naïve and primed pluripotency in an opposing manner. Ribosome footprint (Ribo-Seq) was measured from mouse embryonic stem cells and mouse embriod bodies, in WT and Mettl3-KO cell lines.
2014-11-02 | E-GEOD-61996 | biostudies-arrayexpress
Project description:Gene regulatory elements such as enhancers have profound effects on cellular function, health and disease. Our understanding of mammalian enhancer function is limited by the lack of technology that would allow for a rapid and thorough test of their cell type-specific function. Here we describe a novel Cas9-effector system that enables rapid testing and functional annotation of native enhancers in embryonic stem cells. V6.5 mouse ES cells according to the protocol as described
2015-03-03 | E-GEOD-64058 | biostudies-arrayexpress
Project description:Many large noncoding RNAs (lncRNAs) regulate chromatin, but the mechanisms by which they localize to genomic targets remain unexplored. Here we investigate the localization mechanisms of Xist during X-chromosome inactivation (XCI), a paradigm of lncRNA-mediated chromatin regulation. During the maintenance of XCI, Xist binds broadly across the X-chromosome. During initiation of XCI, Xist initially transfers to distal regions across the X-chromosome that are not defined by specific sequences. Instead, Xist identifies these regions by exploiting the three-dimensional conformation of the X-chromosome. Xist initially accumulates on the periphery of actively transcribed regions and requires its silencing domain to spread across active regions. This suggests a model where Xist coats the entire X-chromosome by searching in three dimensions, modifying chromosome structure, and spreading to newly accessible locations. We examined the genomic localization of the Xist lncRNA using RNA Antisense Purification (RAP) in multiple cell contexts: 1) differentiated female cells (MLFs); 2) a time-course of Xist localization in male embryonic stem (ES) cells where the endogenous Xist promoter is replaced by a tet-inducible one (pSM33); 3) a time-course of Xist localization in differentiating female ES cells (F1 2-1); and 4) wild-type (delXF6) and A-repeat deletion (delSXC9) Xist transgenes incorporated into the Hprt locus under the control of a tet-inducible promoter.
2013-05-30 | E-GEOD-46918 | biostudies-arrayexpress
Project description:Evaluation of CNVs acquired in re-derived embryonic stem cells during targeting or Flp-mediated transgene integration when propagated in N2B27 + 2i + LIF.
2013-11-01 | E-MEXP-3998 | biostudies-arrayexpress